Mixed lymphocyte cultures (MLC) have been studied in which the responding and stimulating strains differed at discrete genetic regions. Differences limited to the D region of the major histocompatibility complex were sufficient to produce both proliferation and the generation of cell-mediated cytotoxicity in vitro. Included in these experiments were combinations differing only at the site of a point mutation in the D region; this difference was also adequate to generate strong responses.